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The selective estrogen receptor modifier tamoxifen (TAM) is widely used for the treatment of women with estrogen receptor positive (ER+ ) breast cancer. Endoxifen (ENDX) is a potent, active metabolite of TAM and is important for TAM's clinical activity. While multiple papers have been published regarding TAM metabolism, few studies have examined or quantified the metabolism of ENDX. To quantify ENDX and its metabolites in patient plasma samples, we have developed and validated a rapid, sensitive, and specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantitative determination of the E- and Z-isomers of ENDX (0.5-500 ng/ml) and the ENDX metabolites norendoxifen (1-500 and 0.5-500 ng/ml E and Z, respectfully), ENDX catechol (3.075-307.5 and 1.92-192 ng/ml E and Z, respectfully), 4'-hydroxy ENDX (0.33-166.5 and 0.33-333.5 ng/ml E and Z, respectfully), ENDX methoxycatechol (0.3-300 and 0.2-200 ng/ml E and Z, respectfully), and ENDX glucuronide (2-200 and 3-300 ng/ml E and Z, respectfully) in human plasma. Chromatographic separation was accomplished on a HSS T3 precolumn attached to an Poroshell 120 EC-C18 analytical column using 0.1 % formic acid/water and 0.1 % formic acid/methanol as eluents followed by MS/MS detection. The analytical run time was 6.5 min. Standard curves were linear (R 2  ≥ 0.98) over the concentration ranges. The intra- and inter-day precision and accuracy, determined at high-, middle-, and low-quality control concentrations for all analytes, were within the acceptable range of 85 % and 115 %. The average percent recoveries were all above 90 %. The method was successfully applied to clinical plasma samples from a Phase I study of daily oral Z-ENDX., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Joel Reid reports financial support and equipment, drugs, or supplies were provided by National Cancer Institute. Matthew Goetz reports a relationship with Lilly that includes: consulting or advisory. Matthew Goetz reports a relationship with Eagle Pharmaceutical Inc that includes: consulting or advisory. Matthew Goetz reports a relationship with Biovica that includes: consulting or advisory. Matthew Goetz reports a relationship with Novartis that includes: consulting or advisory. Matthew Goetz reports a relationship with Pfizer Inc that includes: consulting or advisory. Matthew Goetz reports a relationship with Sermonix that includes: consulting or advisory. Matthew Goetz reports a relationship with AstraZeneca that includes: consulting or advisory. Matthew Goetz reports a relationship with Blueprint Medicines that includes: consulting or advisory. Matthew Goetz reports a relationship with Biotheranostics Inc that includes: consulting or advisory., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Artifacts due to metabolite extraction, derivatization, and detection techniques can result in aberrant observations that are not accurate representations of actual cell metabolism. Here, we show that α-ketoglutarate (α-KG) is reductively aminated to glutamate in methanol:water metabolite extracts, which introduces an artifact into metabolomics studies. We also identify pyridoxamine and urea as amine donors for α-KG to produce glutamate in methanol:water buffer in vitro, and we demonstrate that the addition of ninhydrin to the methanol:water buffer suppresses the reductive amination of α-KG to glutamate in vitro and in metabolite extracts. Finally, we calculate that glutamate levels have been overestimated by 10-50%, depending on cell line, due to α-KG reductive amination. These findings suggest that precautions to account for α-KG reductive amination should be taken for the accurate quantification of glutamate in metabolomics studies., Competing Interests: Declaration of Competing Interest The authors declare no competing interests., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Thiamine deficiency is common in parts of Asia and causes beriberi. Pharmacokinetics of thiamine in deficient populations are unknown., Objective: We characterized thiamine pharmacokinetics in Cambodian mothers and their breastfed infants., Design: Total plasma thiamine, whole-blood thiamine diphosphate (TDP), and breast milk total thiamine were measured in 16 healthy Cambodian mothers and their infants before and after mothers received oral thiamine hydrochloride (100 mg for 5 d). Assays were also performed in 16 healthy American mothers., Results: On day 1, Cambodian mothers were thiamine deficient, with median (range) total plasma thiamine and TDP concentrations of 2.4 nmol/L (0-4.4 nmol/L) and 58.0 nmol/L (27-98 nmol/L), respectively. After a single oral dose, the mean ± SD maximal concentration of thiamine and net area under the thiamine concentration-time curve were 73.4 ± 45.6 nmol/L and 465 ± 241 h · nmol ∙ L⁻¹. Day 6 median maternal total plasma thiamine and TDP concentrations were normal [18.6 nmol/L (13.4-25.3 nmol/L) and 76.5 nmol/L (48-107 nmol/L), respectively; P ≤ 0.001 compared with day 1]. Median Cambodian total breast milk thiamine concentration increased from 180 nmol/L (85-359 nmol/L) on day 1 to 403 nmol/L (314-415 nmol/L) on day 2 and 503 nmol/L (360-808 nmol/L) on day 6; the corresponding American breast milk value was 500 nmol/L (114-622 nmol/L). Median Cambodian infant total plasma thiamine and TDP concentrations increased from 3.0 nmol/L (0-7.3 nmol/L) and 38.5 nmol/L (23-57 nmol/L), respectively, on day 1 to 5.6 nmol/L (0-9.7 nmol/L) and 45.5 nmol/L (32-70 nmol/L), respectively, on day 6., Conclusions: Thiamine-deficient Cambodian mothers effectively absorb oral thiamine, with sharp increases in breast milk thiamine concentrations, but their breastfed infants remain thiamine deficient after 5 d of maternal supplementation. Longer-term maternal supplementation may be necessary to correct thiamine deficiency in breastfed infants. This trial was registered at clinicaltrials.gov as NCT01864057.

Congenital abnormalities of the internal carotid artery (ICA) are infrequent and can be associated with aberrations of the Circle of Willis. A 47-year-old gentleman presented with transient neurological symptoms and cerebral infarction and carotid Doppler showed a stenotic right ICA. Subsequent computed tomographic angiography showed a hypoplastic ICA with a low-lying bifurcation at the C6 level and aplasia of the anterior communicating artery. This patient was commenced on aggressive medical therapy and at 7-month follow-up was symptom-free. This case report highlights the need for a centralized registry with long-term follow-up data in order to identify optimal management., (Copyright © 2012 Society for Vascular Surgery. Published by Mosby, Inc. All rights reserved.)

This study examines correlates of hoarding behaviors among nonclinical elderly adults, focusing upon hoarding cognitions, obsessive-compulsive symptoms, and measures of general psychopathology. Two hundred and sixty-nine adults aged 56-93 years (M=72.49 years) completed the Saving Inventory-Revised, Savings Cognitions Inventory-Revised, Obsessive Compulsive Inventory-Revised, Social Interaction Anxiety Scale, Penn State Worry Questionnaire, and Beck Depression Inventory. Our geriatric sample evidenced significantly greater severity of hoarding behaviors than did a younger community sample (24-72 years; M=44.4 years; Frost, Steketee, & Grishman, 2004). Within our sample, moderate correlations were found between hoarding behaviors and hoarding cognitions; however, controlling for obsessive-compulsive and depressive symptoms resulted in reduced-magnitude associations. As well, relationships between obsessive-compulsive symptoms and both hoarding behaviors and hoarding cognitions were generally strong, with most relationships diminishing after controlling for depressive symptoms. Associations between hoarding symptoms and symptoms of social anxiety, general worry, and depression were generally moderate. A significant regression model showed depressive symptoms explaining the most unique variance in hoarding behaviors. Findings confirm a relatively greater severity of hoarding behaviors in older adults (as compared to younger adults) and suggest that related psychopathology plays a critical role in hoarding expression among older adults. As well, the current study contributes to the ongoing investigation of the diagnostic categorization of compulsive hoarding., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The current study examined factors associated with obsessive-compulsive disorder (OCD) related functional impairment among 99 youth with OCD. A trained evaluator administered the Children's Yale-Brown Obsessive-Compulsive Scale, items assessing family accommodation, and a version of the Brown Assessment of Beliefs Scale that was modified for children. Youth completed the Child Obsessive-Compulsive Impact Scale-Child Version, Obsessive-Compulsive Inventory-Child Version, Multidimensional Anxiety Scale for Children, and Children's Depression Inventory-Short Form. The child's parent completed the Child Obsessive-Compulsive Impact Scale-Parent Version. Results indicated that OCD symptom severity, depressive symptoms, and family accommodation were directly related to impairment, while insight was inversely related to functional impairment. Insight, family accommodation, and depressive symptoms predicted parent- and/or child-rated functional impairment above and beyond OCD symptom severity. Among symptom dimensions, contamination/cleaning and aggressive/checking symptoms were the only dimensions significantly associated with impairment. Assessment and treatment implications are discussed; specifically, we highlight how the variables of interest may impact clinical presentation and treatment course., (Copyright 2009 Elsevier Ltd. All rights reserved.)

Purpose: To describe the pharmacokinetics of 0.5 mg of intravitreal ranibizumab (Lucentis) and to compare it with that of 1.25 mg of intravitreal bevacizumab (Avastin), using the same rabbit model., Design: Experimental animal study., Participants: Twenty-eight Dutch-belted rabbits., Methods: One eye of each of 20 rabbits was injected with 0.5 mg of intravitreal ranibizumab. Both eyes of each of 4 rabbits were enucleated at days 1, 3, 8, 15, and 29. Ranibizumab concentrations were measured in aqueous fluid, whole vitreous, and serum. A further 8 rabbits were used to measure serum and fellow ranibizumab at additional time points of 3 and 8 hours., Main Outcome Measures: Ranibizumab concentrations in the aqueous, vitreous, and serum., Results: Although vitreous concentrations of ranibizumab declined in a monoexponential fashion with a half-life of 2.88 days, concentrations of >0.1 microg/ml ranibizumab were maintained in the vitreous humor for 29 days. Ranibizumab concentrations in the aqueous humor of the injected eye reached a peak concentration of 17.9 microg/ml, 3 days after drug administration. Elimination of ranibizumab from the aqueous humor paralleled that found in the vitreous humor, with a half-life value of 2.84 days. No ranibizumab was detected in the serum or the fellow eye., Conclusion: In the rabbit, the vitreous half-life of 0.5-mg intravitreal ranibizumab is 2.88 days, shorter than the half-life of 1.25-mg intravitreal bevacizumab of 4.32 days. No ranibizumab was detected in the serum or the fellow uninjected eye; whereas small amounts of intravitreal bevacizumab have been detected in the serum and fellow uninjected eye.

Purpose: To describe the pharmacokinetics of 1.25 mg of intravitreal bevacizumab (Avastin)., Design: Experimental animal study., Participants: Twenty Dutch-belted rabbits., Methods: One eye of each of 20 rabbits was injected with 1.25 mg of intravitreal bevacizumab. Both eyes of each of 4 rabbits were enucleated at days 1, 3, 8, 15, and 29. Bevacizumab concentrations were measured in aqueous fluid, whole vitreous, and serum., Main Outcome Measures: Bevacizumab concentrations in the aqueous, vitreous, and serum., Results: Whereas vitreous concentrations of bevacizumab declined in a monoexponential fashion with a half-life of 4.32 days, concentrations of >10 microg/ml bevacizumab were maintained in the vitreous humor for 30 days. Bevacizumab concentrations in the aqueous humor of the injected eye reached a peak concentration of 37.7 microg/ml 3 days after drug administration. A maximum serum concentration of 3.3 mug/ml was achieved 8 days after intravitreal injection and the concentration fell below 1 microg/ml 29 days after injection. Elimination of bevacizumab from the aqueous humor and serum paralleled that found in the vitreous humor, with half-life values of 4.88 days and 6.86 days, respectively. Very low concentrations of bevacizumab were detected in the fellow uninjected eye. Concentrations of bevacizumab in the vitreous of the fellow eye varied incrementally, from 0.35 ng/ml at 1 day to 11.17 ng/ml at 4 weeks. Concentrations of bevacizumab in the aqueous humor of the fellow eye reached their peak at 1 week, at 29.4 ng/ml, and declined to 4.56 ng/ml at 4 weeks., Conclusion: The vitreous half-life of 1.25 mg intravitreal bevacizumab is 4.32 days in a rabbit eye. Very small amounts of bevacizumab were detected in the serum and in the fellow uninjected eye.

The dimeric pyrrolobenzodiazepine SJG-136 (NSC 694501) has potent in vitro cytotoxicity and in vivo antitumor activity. SJG-136 binds in the minor groove of DNA and produces G-G interstrand cross-links via reactive N(10)-C(11)/N(10')-C(ll') imine/carbinolamine moieties. We have developed a sensitive, specific liquid chromatography tandem mass spectrometry (LC/MS/MS) method for the quantitative determination of SJG-136 in plasma. SJG-136 was isolated by solid phase extraction through a C8 column, reverse-phase HPLC separation was accomplished on a C18 column with isocratic elution and MS/MS detection, monitoring the m/z 557-m/z 476 transition after electrospray ionization. The linear range and lower limit of quantitation from plasma standard curves were 2.8-1800 nM, and 5 nM, respectively. SJG-136 plasma protein binding was species-dependent. Values of the unbound fraction in human, rat and mouse were 25%, 16.2% and <1%, respectively. Protein binding was saturable in dog plasma where the unbound fraction increased from 10.8% to 22.3% over a 22-720 nM concentration range. SJG-136 pharmacokinetics after a single intravenous dose were best fit to a two-compartment open model with elimination half-life and plasma clearance values of 97 min and 6.1 mL/min/kg, respectively. SJG-136 did not accumulate in plasma following intravenous administration of 1.0 microg/kg doses for five consecutive days.

Dacarbazine (DTIC) is a prodrug that is clinically effective in the treatment of Hodgkin's disease, melanoma and soft tissue sarcoma. To better characterize the clinical pharmacology of parent drug and reactive metabolites, a reversed-phase HPLC method with UV detection was developed for simultaneous determination of dacarbazine and the metabolites 5-(3-hydroxymethyl-3-methyl-1-triazeno)imidazole-4-carboxamide (HMMTIC) and 5-(3-methyl-1-triazeno)imidazole-4-carboxamide (MTIC). Chromatographic separation was achieved with a Zorbax SB-CN column and with a mobile phase of 80% 50 mM ammonium phosphate, pH 6.5, 20% methanol and 0.1% triethylamine. HMMTIC, MTIC and DTIC were extracted from plasma with methanol precipitation of the proteins. Recovery of DTIC and the metabolites from whole blood was greater than 92%. Rapid processing of whole blood, methanol extraction and storage at -70 degrees C substantially increased the stability of HMMTIC and MTIC from less than 15 min to 3 days. Precision for HMMTIC, MTIC and DTIC ranged from 3.7 to 16.3% relative standard deviation. The accuracy ranged from 101 to 114% for all three analytes. The validated assay was used to determine the pharmacokinetic data for dacarbazine and its active metabolites for human patients with recurrent glioma receiving DTIC intravenously.

The regulation of membrane trafficking is thought to be predominantly under the control of agonist-receptor transduction pathways. In the present study, osmomechanical stress due to swelling, a condition often accompanying cell activation, was shown to induce multiple membrane trafficking pathways in polarized absorptive epithelial cells in the absence of agonists. Osmomechanical stress activated rapidly (seconds) pathways of calcium-dependent membrane insertion into the basolateral domain, pathways of calcium-independent membrane retrieval from the basolateral domain, and a novel pathway of transcytosis (transcellular) between basolateral and apical cell domains. These pathways appear to underlie the transfer and regulation of transport proteins amongst cell compartments. This broad affect of osmomechanical stress on trafficking pathways may reflect a global mechanism for redistribution of transport proteins and other membrane components amongst cell compartments during states of mechanical stress., (Copyright 2000 Academic Press.)

It has recently been shown that rabbit kidney epithelial cells (proximal tubule) possess two dihydropyridine (DHP)-sensitive calcium entry channels (1, 2). To evaluate the properties of the DHP binding sites, the binding of the DHP, [3H]PN200-110, was studied in epithelial cell membrane fractions (proximal tubule) of rabbit kidney cortex. High-affinity binding sites for the DHP were observed in both basolateral and apical membranes and in a membrane microsomal preparation from rabbit primary cultures of proximal tubule cells (cultured PT). In an extended analysis of the basolateral membrane preparation, two high-affinity binding sites were evident with binding dissociation constants, Kd, of 0.005 and 0.75 nM. The Kd values are similar to that observed for L-type calcium channel alpha1-subunits. Using a homology-based cloning strategy, a 388-base fragment of an alpha1-subunit was cloned from RNA isolated from rabbit cultured PT cells and freshly isolated proximal tubules and found to encode a protein identical to the cardiac form of the L-type alpha1-subunit (alpha1C-subunit). It is concluded that renal epithelial cells express high-affinity dihydropyridine receptors and that the receptors may be components of plasma membrane calcium channels, including L-type calcium channels, that control calcium entry in these cells.

The ultrasonic absorption of sickle cells was compared with the morphological changes in these cells are produced by varying oxygen tensions and pH. A decrease in oxygen tension caused increased sickling and an increase in the ultrasonic absorption of the blood. An increase in pH reduced the ultrasonic absorption and decreased sickling. These findings indicate that ultrasonic absorption can detect changes in the aggregation state of sickle cell hemoglobin in situ within the red cell and that the method may be used to monitor the degree of sickling under various environmental conditions.